1. Field of the Invention
The present invention relates generally to the field of cancer therapy and prophylaxis. More particularly, the present invention provides growth factor antagonists which inhibit the growth of cancers including tumors and pre-cancerous tissue. Even more particularly, the present invention is directed to antagonists of vascular endothelial growth factor-B and their use to inhibit the growth of cancer including tumor tissue and pre-cancerous tissue.
2. Description of the Prior Art
Bibliographic details of the publications referred to in this specification are also collected at the end of the description.
Reference to any prior art is not, and should not be taken as an acknowledgement or any form of suggestion that this prior art forms part of the common general knowledge in any country.
The development of blood vessels and of vascular supply is a fundamental requirement for organ development and differentiation during embryogenesis, as well as for normal postnatal physiological processes such as wound healing, tissue and organ regeneration, and cyclical growth of the corpus luteum and endometrium (Folkman & Klagsbrun, Science, 235(4787): 442-447, 1987; Klagsbrun & D'Amore., Ann. Rev. Physiol. 53:217-239, 1991; Carmeliet et al., Nature 380:435-439, 1996; Ferrara et al., Nature 380:439-442, 1996). The growth and maturation of new vessels (angiogenesis) is a highly complex and coordinated process and requires the sequential activation of a series of receptors by numerous ligands (Yancopoulos et al., Nature 407:242-248, 2000; Ferrara and Alitalo, Nature Medicine 5:1359-1364, 1999; Carmeliet, Nature 407:249-257, 2000). Vascular endothelial growth factor (VEGF or VEGF-A) is arguably the most thoroughly characterized of these ligands and VEGF-A signaling appears to represent a critical rate-limiting step in the process (Ferrara et al., Nature Medicine 9:669-676, 2003).
In addition to normal physiological processes, the pathological growth of tumors is also known to be dependent on the degree of new blood vessel formation in the tumor bed (Carmeliet et al., 2000 supra; Folkman, Nature Medicine 1:27-31, 1995; Hanahan & Folkman, Cell 86:353-364, 1996). VEGF-A mRNA is upregulated in many human tumors and VEGF-A appears to be an important angiogenic factor frequently utilized by tumors to switch on blood vessel growth (Dvorak et al., Semin Perinatol 24:75-78, 2000; Ferrara & Alitalo, 1999 supra; Yancopoulos, 2000 supra; Benjamin & Keshet, Proc Natl Acad Sci USA 94:8761-8766, 1997; Ferrara and Davis-Smyth, Endocr. Rev 18:4-25, 1997). VEGF-A also increases vascular permeability, and this is thought to be important for tumor invasion and metastasis (Dvorak et at., Curr Top Microbiol Immunol 237:97-132, 1999). As a result there has been a significant effort towards the development of agents that target angiogenic factors such as VEGF-A in order to inhibit tumor growth (Ferrara et al., 2003 supra). One such agent is bevacizumab, a humanized mouse monoclonal antibody that binds to, and inhibits the activity of, VEGF-A. Bevacizumab (Avastin) has recently been approved by the FDA for the treatment of colorectal cancer.
VEGF-A is now recognized as the founding member of a family of structurally related molecules. The ‘VEGF family’ comprises six members including prototype VEGF-A, placenta growth factor (PLGF), VEGF-B, VEGF-C, VEGF-D and VEGF-E (Eriksson & Alitalo, Curr Top Microbiol Immunol 237:41-57, 1999). The biological functions of the VEGF family are mediated by the differential activation of at least three structurally homologous tyrosine kinase receptors, VEGFR-1/Flt-1, VEGFR-2/Flk-1/KDR and VEGFR-3/Flt-4. VEGF-A, VEGF-B and PLGF also bind to the non-tyrosine kinase receptors neuropilin-1 and -2, Soker et al., Cell 92:735-45, 1998; Neufeld et al., Trends Cardiovasc Med. 12:13-19, 2002). According to their receptor binding patterns, the VEGF family can be divided into three subgroups: (1) VEGF-A, which binds to VEGFR-1 and VEGFR-2; (2) PLGF and VEGF-B, which bind only to VEGFR-1 and; (3) VEGF-C and VEGF-D, which interact with both VEGFR-2 and VEGFR-3 (Ferrara & Alitalo, 1999 supra; Ferrara et al., 2003 supra).
As noted above, VEGF-A is the most thoroughly characterized member of the VEGF family and an accumulation of evidence has led to the conclusion that VEGFR-2 is the major mediator of VEGF-A associated biological activities such as endothelial cell proliferation, migration and survival, angiogenesis and vascular permeability (Ferrara et al., 2003 supra). In addition to VEGFR-2, VEGF-C and -D also bind to, and activate, VEGFR-3. VEGFR-3 is expressed primarily on lymphatic endothelial cells and VEGF-C and -D are thought to be key regulators of lymphatic angiogenesis [or lymphangiogenesis] (Makinen et al., Nature Medicine 7:199-205, 2001; Skobe et al., Nature Medicine 7:192-8, 2001; Stacker et al., Nature Medicine 7:186-91, 2001). In contrast to VEGF-A, -C and -D and the downstream effects of signaling through VEGFR-2 or -3, the precise role of VEGF-B and signaling through VEGFR-1 remains poorly understood.
VEGFR-1 is expressed on a variety of cell types (Clauss et al., J. Biol. Chem. 271:17629-17634, 1996; Wang & Keiser, Circ. Res. 83:832-840, 1998; Niida et al., J. Exp. Med. 190:293-298, 1999) and expression, at least on endothelial cells, is upregulated by hypoxia and a HIF-1α dependent mechanism (Gerber et al., J. Biol. Chem. 272:23659-23667, 1997). However, only weak autophosphorylation of VEGFR-1 is observed in response to VEGF-A, and VEGF-A binding to VEGFR-1 appears not to activate the downstream signals required for key endothelial cell responses such as proliferation and survival (de Vries et al., Science 255:989-991, 1992; Waltenberger et al., J. Biol. Chem. 269:26988-26995, 1994; Keyt et al., J. Biol. Chem. 271:5638-5646, 1996; Rahimi et al., J. Biol. Chem. 275:16986-16992, 2000). The observation that the VEGFR-1 specific ligand, PLGF, enhanced the activity of VEGF-A on endothelial cells suggested that VEGFR-1 might function as a decoy receptor ie. PLGF displaced VEGF-A from VEGFR-1 making it available to bind to, and signal through VEGFR-2 (Park et al., J Biol. Chem. 269:25646-25654, 1994). In vivo data from genetically modified mice further suggested a non-signaling decoy role for VEGFR-1. VEGFR-1−/− mice died in utero between days 8.5 and 9.5 and although endothelial cells developed, they failed to organize into vascular channels (Fong et al., Development 126:3015-3025, 1999). Lethality was attributed to excessive angioblast proliferation and this in turn, was attributed to enhanced VEGF-A action (Fong et al., 1999 supra). The observation that mice expressing VEGFR-1 lacking the kinase domain were healthy and showed no overt defect in vascular development provided further support for the decoy hypothesis, as the truncated receptor could still bind VEGF-A, but not transmit intracellular signals (Hiratsuka et al., Proc. Natl. Acad. Sci. USA 4:9349-9354, 1998).
Analysis of VEGF-B−/− mice has also failed to resolve the confusion surrounding the precise physiological (and pathological) role of VEGFR-1 specific ligands and VEGFR-1 signaling. In contrast to VEGF-A−/− mice, VEGF-B−/− mice display no overt defects in vascular development and are healthy and fertile (Bellomo et al., Circ. Res. 86:E29-E35, 2000). In one report the hearts of VEGF-B−/− mice were reduced in size and the response to coronary occlusion and myocardial recovery from ischemia were compromised (Bellomo et al 2000, supra). Although heart morphology appeared normal, the authors concluded that VEGF-B is essential for the establishment of a fully functional coronary vasculature. In contrast, a second report describing VEGF-B−/− mice reported only a minor atrial conduction defect (Aase et al., Circulation 104:358-364, 2001).
As a result of the confusion surrounding the role for VEGFR-1 and VEGFR-1-specific ligands in the regulation of blood vessel formation, the potential of VEGF-B as a therapeutic target for inhibition of tumor growth and metastasis is unclear. Although VEGF-B has been shown, along with other factors, to be expressed in a variety of tumors (Salven et al., Am J Pathol, 153:103-108, 1998), evidence of upregulation is limited (Li et al., Growth Factors 19:49-59, 2001) and there have been no reports of the efficacy of VEGF-B specific antagonists in xenograft or other relevant animal models. In fact, the potential of VEGF-B (and PLGF) is further confused by the disclosure by Cao et al in International PCT Publication No. WO03/62788, which suggests that increasing VEGF-B expression inhibits VEGF-A induced angiogenesis, and thus represents a potential approach to the treatment of diseases caused by VEGF-A activity and VEGF-A induced angiogenesis.
In accordance with the present invention, it has been surprisingly determined that antagonists of VEGF-B are useful in reducing the growth and development of cancer including tumor tissue.